Functional characterization of AtRUG8, a protein that modulates plasmodesmal movement of macromolecules, and of three 3 closely related proteins

• Applicant: Professor Dr. Norbert Sauer
• Subject Area: Plant Physiology
• Term: from 2009 to 2013
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 121466635

We have shown that soluble GFP that is synthesized in Arabidopsis or tobacco (Nicotiana tabacum) companion cells (CCs) under the control of the Arabidopsis AtSUC2 (SUCROSE TRANSPORTER 2) promoter moves from CCs into sieve elements (SEs), traffics with the photoassimilates from source to sink, and is eventually symplastically unloaded into different terminal sink tissues, such as anthers, young leaves or root tips. Screening of more than 7,000 T-DNA insertion lines that we generated in plants with an AtSUC2-promoter::GFP transgene, identified eight mutants with significantly reduced cell-to-cell mobility of GFP. These mutants were named “reduced unloading of GFP” (rug). So far, one of these mutants, rug8, has been partly characterized on the molecular level. In two independent mutants (rug8-1 & rug8-2) the functionally uncharacterized gene At1g17930 was shown to be responsible for the observed phenotype, and complementation with the wild type sequence restored the mobility of soluble GFP. RUG8 has an N-terminal domain of unknown function (DUF1723) and a predicted Cterminal nuclear localization sequence (NLS). In fact, analyses of the subcellular localization confirmed that RUG8 localizes to the nucleus and a yeast 2-hybrid screen revealed interaction of RUG8 with the AP2/ERF-type transcription factor ERF38, an Arabidopsis protein that regulates the expression of genes involved in cell wall thickening. In the proposed project we will study the physiological role of the RUG8 protein and the effect of rug8 mutations on the size exclusion limit (SEL) of plasmodesmata. Moreover, we will screen for additional RUG8 interactors, search for altered gene expression in rug8 mutants, and study the mode of interaction between RUG8 and ERF38. Eventually we will extend these analyses to the three closely related Arabidopsis genes At1g48120, At2g04865 and At2g25010.

• DFG Programme: Research Grants